Electron-transfer reactions play a key role in oxidative phosphorylation and respiration, drug and carcinogen metabolism and activation, immune response, collagen synthesis, and intermediary metabolism. Our aim is to probe some of the factors that control the rates of long-range electron-transfer processes in metalloenzymes: driving force, temperature, site-to-site distance, and the nature of the protein medium and the electron acceptor site. We intend to study intramolecular long-range electron-transfer reactions in ruthenium (II/III) derivatives of three structurally well-characterized copper proteins: P. aeruginosa azurin, bovine erythrocyte superoxide dismutase, and Cu(II)-substituted horse liver alcohol dehydrogenase. During the next three years we propose to: (1) prepare highly purified and well-characterized samples of the above three proteins labelled at selected surface histidine residues with ruthenium (II/III) reagents: (2) determine by flash photolysis experiments the rates of electron transfer from the surface-bound ruthenium (II) reagents to the copper (II) sites in the interiors of these proteins, thereby providing important kinetic information relating directly to electron tunnelling through proteins; and (3) measure the pH and temperature dependences of the reduction potentials in the native and modified copper proteins by spectroelectrochemical techniques, which will give us a detailed picture of the factors that control the thermodynamics of these long-range electron-transfer reactions.